The objective of this proposal is to define the structure-function relationships of the human factor VIII/von Willebrand (FVIII/vWF) protein(s). The following will be done: 1) experiments to determine if the "dissociable" FVIII procoagulant activity protein is a proteolytic derivative of a single large parent molecule with a subunit structure linked by disulfide bonds and potentially both FVIII procoagulant and vWF activities; 2) studies to define the molecular basis for vWF activity. These two objectives require that each protein(s) associated with either FVIII procoagulant or vWF activity be purified and characterized biochemically, i.e., by sedimentation equilibrium molecular weights, subunit structure, amino-terminal analyses, amino acid compositions, carbohydrate analyses, cyanogen bromide cleavage patterns and by digestion patterns by various proteolytic enzymes. The effect of removing successive carbohydrate groups from FVIII/vWF on its vWF activity, defined by ristocetin-platelet aggregation, and on its in vivo plasma survival will be studied. Simultaneously, binding and inhibition of binding of each carbohydrate-modified FVIII/vWF species to the hepatic receptor for terminal galactosyl will be studied. The ability of sialyltransferases to re-sialylate asialo-FVIII/vWF will be quantitated by: 1) the amount of sialic acid reattached; 2) the amount of restored vWF activity; and 3) the extent of binding to the hepatic receptor. Finally, the isolated cyanogen bromide and plasmin-cleaved fragments of FVIII/vWF will be tested for support of ristocetin-induced platelet aggregation; those which do will be examined for carbohydrate and how it relates to their having vWF activity. Preliminary studies of one plasmin-cleavage fragment of FVIII/vWF show that it has carbohydrate, supports ristocetin-platelet aggregation and may contain a region of the molecule important to FVIII procoagulant function. These studies should expand our insights into the role of these two clotting activities in normal blood coagulation, abnormal states of hemostasis and thrombotic disorders.